Balla Helga, Borsodi Kinga, Őrsy Petra, Horváth Béla, Molnár Péter József, Lénárt Ádám, Kosztelnik Mónika, Ruisanchez Éva, Wess Jürgen, Offermanns Stefan, Nyirády Péter, Benyó Zoltán
Institute of Translational Medicine, Semmelweis University, Budapest, Hungary.
Department of Urology, Semmelweis University, Budapest, Hungary.
Am J Physiol Renal Physiol. 2023 Nov 1;325(5):F618-F628. doi: 10.1152/ajprenal.00261.2022. Epub 2023 Sep 7.
Acetylcholine plays an essential role in the regulation of detrusor muscle contractions, and antimuscarinics are widely used in the management of overactive bladder syndrome. However, several adverse effects limit their application and patients' compliance. Thus, this study aimed to further analyze the signal transduction of M and M receptors in the murine urinary bladder to eventually find more specific therapeutic targets. Experiments were performed on adult male wild-type, M, M, M/M, or Gα knockout (KO), and pertussis toxin (PTX)-treated mice. Contraction force and RhoA activity were measured in the urinary bladder smooth muscle (UBSM). Our results indicate that carbamoylcholine (CCh)-induced contractions were associated with increased activity of RhoA and were reduced in the presence of the Rho-associated kinase (ROCK) inhibitor Y-27632 in UBSM. CCh-evoked contractile responses and RhoA activation were markedly reduced in detrusor strips lacking either M or M receptors and abolished in M/M KO mice. Inhibition of Gα-coupled signaling by PTX treatment shifted the concentration-response curve of CCh to the right and diminished RhoA activation. CCh-induced contractile responses were markedly decreased in Gα KO mice; however, RhoA activation was unaffected. In conclusion, cholinergic detrusor contraction and RhoA activation are mediated by both M and M receptors. Furthermore, whereas both Gα and Gα proteins mediate UBSM contraction, the activation at the RhoA-ROCK pathway appears to be linked specifically to Gα. These findings may aid the identification of more specific therapeutic targets for bladder dysfunctions. Muscarinic acetylcholine receptors are of utmost importance in physiological regulation of micturition and also in the development of voiding disorders. We demonstrate that the RhoA-Rho-associated kinase (ROCK) pathway plays a crucial role in contractions induced by cholinergic stimulation in detrusor muscle. Activation of RhoA is mediated by both M and M receptors as well as by G but not G proteins. The G-RhoA-ROCK pathway may provide a novel therapeutic target for overactive voiding disorders.
乙酰胆碱在逼尿肌收缩调节中起重要作用，抗毒蕈碱药物广泛用于治疗膀胱过度活动症。然而，一些不良反应限制了它们的应用及患者的依从性。因此，本研究旨在进一步分析小鼠膀胱中M和M受体的信号转导，最终找到更具特异性的治疗靶点。对成年雄性野生型、M、M、M/M或Gα基因敲除（KO）以及经百日咳毒素（PTX）处理的小鼠进行实验。测量膀胱平滑肌（UBSM）的收缩力和RhoA活性。我们的结果表明，氨甲酰胆碱（CCh）诱导的收缩与RhoA活性增加相关，并且在UBSM中存在Rho相关激酶（ROCK）抑制剂Y-27632时收缩力降低。在缺乏M或M受体的逼尿肌条中，CCh诱发的收缩反应和RhoA激活明显降低，而在M/M KO小鼠中则完全消失。PTX处理抑制Gα偶联信号转导使CCh的浓度-反应曲线右移，并减少RhoA激活。CCh诱导的收缩反应在Gα KO小鼠中明显降低；然而，RhoA激活未受影响。总之，胆碱能逼尿肌收缩和RhoA激活由M和M受体介导。此外，虽然Gα和Gα蛋白均介导UBSM收缩，但RhoA-ROCK途径的激活似乎与Gα特异性相关。这些发现可能有助于确定膀胱功能障碍更具特异性的治疗靶点。毒蕈碱型乙酰胆碱受体在排尿的生理调节以及排尿障碍的发生发展中至关重要。我们证明RhoA-Rho相关激酶（ROCK）途径在胆碱能刺激诱导的逼尿肌收缩中起关键作用。RhoA的激活由M和M受体以及G介导，但不由G蛋白介导。G-RhoA-ROCK途径可能为膀胱过度活动症提供新的治疗靶点。
